Multiple N-methytransferases for aromatic alkylamines in brain

Abstract

This report describes our studies of the enzymatic N-methylation of tryptamine and β-phenylethylamine by fractions of homogenized rat brain with either 5-methyltetrahydrofolic acid (5-MTHF) or S-adenosyl-L-methionine (SAM) as the methyl donor. We found the pH optimum between 6.5 and 7.0 in reaction involving 5-MTHF and either substrate. (We confirmed reports that reactions involving SAM have a pH optimum of 7.9 with either substrate). Enzymatic activity in the presence of 5-MTHF was linear with time and protein concentration. Measures of enzymatic activity in the 100,000 x g supernate of whole rat brain homogenerate were enriched 15–25 fold after Sephadex G-100 fractionation of the 40–50% ammonium sulfate precipitate. Dialysis enhanced the enzyme activity in virtually all of the fractions. Blanks with tryptamine and boiled enzyme gave unexpectedly high counts (comparable to those without enzyme), suggesting possible non-enzymatic methylation in addition to enzymatic N-methylation. We controlled for this by using blanks of substrate plus boiled enzyme. Radioactive products were isographic with N-monomethyltryptamine and N,N-dimethyl tryptamine on thin layer chromatograms. In the presence of 5-MTHF there is low enzymatic affinity (Km = approximately 10 −3 M) for either substrate, suggesting that enzymatic N-methylation might occur only when amine concentrations are abnormally high. Assays in the presence of each donor in combination with each substrate suggested the possible existence of multiple N-methyltransferases, with one specific for tryptamine and the others non-specific for aromatic alkylamines.